The Promise of Regenerative Medicine

In August 2006 Professor Yamanaka Shin’ya of Kyoto University published a paper reporting his success in producing induced pluripotent stem cells, and in a 2014 clinical study, tissue derived from iPS cells was transplanted to a patient for the first time. While a host of challenges must be overcome, the promise of regenerative therapies appears to be coming within reach.

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Regenerative medicine has been a dream of researchers for years. The ultimate goal of this field is to restore the functions of failing body parts: eyes that have lost sight, limbs that no longer move, hearts on the verge of stopping. Replacing lost or diminished functions with eyeglasses, prosthetic limbs, and artificial organs may be considered regenerative medicine in a broad sense. A breakthrough loomed on the horizon 35 years ago, when scientists succeeded in producing embryonic stem cells from the embryos of mice in 1981. And in 1998 they were able to produce human ES cells.

The adult human body consists of roughly 60 trillion cells, all of which were generated from a single fertilized egg through repeated cell division. Stem cells are capable of replicating themselves through division and differentiating into a variety of cells—an ability called pluripotency. The fertilized egg, in particular, can produce every kind of adult cell.

ES cells are made from early embryos that have undergone only six or seven cell divisions after fertilization. In essence, they are stem cells that can proliferate almost indefinitely without losing their pluripotency. While offering hope in regenerative therapy, ES cells also faced a number of issues, such as the need for technologies to safely induce differentiation in targeted cells and to prevent rejection following transplantation. The biggest barrier, though, was ethical, as stem cells had to be made from human embryos.

As researchers around the world struggled to induce the differentiation of ES cells into various somatic cells, Yamanaka Shin’ya, a professor at Kyoto University, succeeded in producing pluripotent cells by taking an inverse approach.

Winning the Nobel Prize

Examining the 24 genes that are highly expressed in ES cells, Yamanaka ascertained 4 that later came to be known as the Yamanaka factors: Oct3/4, Sox2, Klf4, and c-Myc. Incorporating them into the skin cells of mice using retroviruses as carriers (vectors) resulted in pluripotent stem cells similar to ES cells. Yamanaka named these cells “induced pluripotent stem cells,” or iPS cells. The “i” was lowercased in the hope of repeating the global success of the iPod, Apple’s portable music player.

The details of the production process and the identity of these four genes were revealed in the August 25, 2006, issue of the US scientific journal Cell. Before iPS cells could be used in treating human patients, however, Yamanaka needed to prove that they could be produced with human rather than mouse cells. While the leap from mouse-derived to human-derived ES cells took 17 years, he succeeded in producing human iPS cells the following year.

A colony of human iPS cells established from fibroblasts. The width of the colony is about 0.5 mm. (Photo courtesy of Yamanaka Shin’ya)

The iPS cells have the potential of changing not just regenerative medicine but the entire face of clinical medicine. By comparing a patient’s diseased tissue with iPS cells produced from the same patient’s reprogrammed cells, for instance, the mechanism of the disease might be unraveled and new drugs developed. Even US President George W. Bush and Pope Benedict XVI, who had been opposed to ES cells on ethical grounds, welcomed the development of iPS cells. In 2012, before his discovery had saved a single human life, Yamanaka was awarded the Nobel Prize in Physiology or Medicine.

The development of human iPS cells signaled the start of a race to realize the dream of regenerative medicine.

The initially developed method of producing iPS cells was plagued by the risk of tumor formation and was highly inefficient. After enhancing cell safety by adjusting the choice of the genes used and the method of production, clinical tests were launched in 2014. On September 12, iPS-cell-derived retinal pigment epithelial sheets prepared from the patient’s skin cells were transplanted to the eye of a woman in her seventies suffering from exudative age-related macular degeneration, an intractable disease. The operation was conducted at the Institute of Biomedical Research and Innovation Hospital in Kobe, Hyōgo Prefecture.

The ongoing tests are being headed by Takahashi Masayo, project leader at the Riken scientific research institute and an eye surgeon. The primary objective of the trial is to evaluate the safety of the treatment; the study will track the engraftment and check for tumor formation over a period of more than four years. No problems have been found in the two years that have elapsed so far. But the second clinical trial, which had been scheduled to begin in 2015, was canceled after researchers found multiple genetic mutations in the iPS cells sourced from the patient’s cells.

Suppressing Immune Rejection and Tumorigenesis

Cultivating iPS cells from the patient’s own cells has the advantage of avoiding immune rejection, which has been a major concern with transplantation. In the Riken trial, though, 11 months were required to cultivate patient-derived iPS cells, differentiate them, and further shape them into retinal sheets, conducting thorough safety checks along the way. The cost also snowballed to several hundred million yen. For the next trial, therefore, Riken will seek to generate retinal pigment epithelial cells for transplantation by differentiating stock iPS cells from Kyoto University’s Center for iPS Cell Research and Application (CiRA). This should allow researchers to resume transplantation in 2017.

Cells fall into one of several human leukocyte antigen (HLA) types, somewhat analogous to ABO blood typing. Just as type O blood can be transfused to individuals of every blood type, certain HLA types have been known not to trigger a rejection response when transplanted. Kyoto University is in the process of stockpiling iPS cells of the most widely applicable HLA types.

Tanks storing the iPS cell stock at the Facility for iPS Cell Therapy (FiT) in CiRA. (Photo courtesy of CiRA)

Tens of thousands of HLA types are thought to exist, and individuals who have inherited the same HLA markers from both parents have what is known as homozygous HLA, which may be expressed, in simplified terms, as AA, BB, or CC. For example, the cells of a person having the AA genotype can be transplanted to a patient having an AB or AC genotype with minimum risk of rejection. Just 75 HLA-homozygous donors (with the 75 most common HLA types) are thought to be enough to cover 80% of the population, while having 140 such donors would boost the coverage rate to 90%. As an iPS cell stock of this size is not too difficult to develop, Kyoto University has enlisted the help of Kyoto University Hospital, the Japanese Red Cross Society, cord blood banks, and other institutions in identifying HLA-homozygous individuals, who are then asked to cooperate as donors.

Using donated cells, Kyoto University is developing a reserve of high-quality iPS cells for treatment, which will be applicable to a projected 30%–50% of the Japanese population by the end of March 2018.

The risk of tumor formation, which has been the largest technological hurdle from the very start of clinical research, has not been eliminated despite improvements to the production method. Researchers seek to induce iPS cells to differentiate into the desired cell type, but if any undifferentiated iPS cells remain, these could turn into tumors. Finding ways to either fully differentiate all of the cells or remove any impurities will be an important key to the success of regenerative therapy using iPS cells.

Paving the Legal Way to Practical Use

The Japanese government is providing indirect support toward the practical use of iPS cells through legal revisions and other means. The Pharmaceutical Affairs Law was amended in November 2014 so that regenerative medical products, such as cell sheets, whose safety has been confirmed in clinical trials will receive government approval as pharmaceutical products as soon as they are presumed effective, albeit with restrictions on to whom they can be sold and for how long. If the product’s efficacy can be fully verified within a certain time period after going on sale, it will be reviewed and formally approved. This is expected to cut the time needed for approval from over 10 years to just 2–3 years.

The revised law is said to be at the cutting edge of pharmaceutical legislation worldwide. Approving regenerative medical products ahead of other countries entails significant risks, but the Japanese government appears ready to shoulder those risks in supporting iPS research.

In the interest of protecting patients, meanwhile, a separate law has been passed to prevent the rampancy of clinics offering regenerative therapy of unproven efficacy and safety at the patient’s own risk and expense.

New Treatments

Research on using iPS cells to treat heart failure, spinal cord injury, Parkinson’s disease, and a variety of other conditions is steadily advancing, gradually moving past the animal experimentation stage and approaching that of clinical studies on human subjects.

Professor Okano Hideyuki, dean of the Keiō University School of Medicine, is working to make treatments using iPS cells available to patients with spinal cord injury. Because such injuries sustained from accidents and trauma become chronic quickly, there is no time to wait for the growth of patient-derived iPS cells. As such, Okano plans to use Kyoto University’s iPS cell stock to produce differentiated neural stem cells, which will then be injected in the first human subject during the 2017–18 fiscal year. He also hopes to apply this technology to the treatment of cerebral infarction.

Professor Sawa Yoshiki of Osaka University is developing a treatment for heart failure in which iPS-cell-derived myocardial cell sheets are grafted onto the heart.

Professor Takahashi Jun of Kyoto University is working on a treatment for Parkinson’s disease, an incurable neurological disorder, that uses dopamine-producing nerve cells derived from iPS cells. To help meet the shortage of donors in an aged society, moreover, Takahashi is conducting a project in collaboration with the Japanese Red Cross Society to manufacture platelets and red blood cells from iPS cells. Other plans include treating cancer by using iPS cell technology to activate immune cells.

Meanwhile, efforts are also being made to grow entire organs from iPS cells. Professor Taniguchi Hideki of Yokohama City University has succeeded in developing a three-dimensional “liver bud” from precursor cells (partially differentiated cells on the way to becoming somatic cells of a particular kind) induced from human iPS cells.

Use of iPS Cells in Drug Development

Along with regenerative therapy, drug development has been regarded as the other major field in which iPS cells can be applied.

Tens of thousands of substances are tested before one is approved as a new drug. It is not unusual for developers to give up on a promising substance due to strong side effects or other factors that emerge during development, and iPS cells could serve as a tool for predicting such negative effects. For instance, liver toxicity can be predicted by using liver cells induced from human iPS cells. Arrhythmia, another potentially fatal side effect, has already become predictable with the aid of iPS-cell-derived myocardial cells.

High expectations are also being placed on iPS cells for the development of drugs for intractable diseases. In 2012 the Ministry of Education, Culture, Sports, Science, and Technology and the Ministry of Health, Labor, and Welfare launched a project for industry-academia collaboration in research on intractable diseases utilizing iPS cells. By producing iPS cells from somatic cells (such as skin and blood cells) donated by patients with intractable diseases and then differentiating them into cells of the affected area, the project aims to elucidate the pathology of these diseases and pave the way for the development of new drugs. Fifty MHLW research teams that study intractable diseases are coordinating with the project, which involves five centers including Kyoto University, and seven pharmaceutical companies are also taking part. Kyoto University and Keiō University have pinpointed potential curatives for six diseases, including fibrodysplasia ossificans progressiva, from among the drugs that are currently used to treat other diseases. They will aim to make these drugs available for practical use after clinical trials and other procedures.

The Japanese government is investing tens of billions of yen toward the practical application of treatments using iPS cells, but issues remain in human resources development and other areas. The Japanese Society for Regenerative Medicine is addressing the shortage of workers by launching an accreditation scheme for certified doctors in regenerative medicine and clinical cell culture engineers and by training technicians skilled in cultivating and processing cells.

A decade has passed since the discovery of iPS cells, and clinical research has begun, but there is still a long way to go before their benefits reach all corners of society. Yamanaka Shin’ya, who pioneered the field, and other scientists around the world are working tirelessly to move research forward while walking the line between safety and speed, and between the risks and benefits—propelled by their faith in the immense potential of iPS cells.

(Originally written in Japanese and published on September 28, 2016. Banner photo: A researcher conducts an experiment in an open laboratory at the Kyoto University Center for iPS Cell Research and Application. Photo courtesy of CiRA, Kyoto University.)